This application is or renewal of grant AM21458. The overall objective of this project is to utilize model cell culture systems to study hormone synthesis by endocrine cells. The hormone-producing cells utilized for these studies include the JEG clonal lines of human choriocarcinoma cells which synthesize the glycoprotein hormone, chorionic gonadotropin as well as another protein hormone, placental lactogen. These cells also produce progesterone and convert appropriate 19-carbon steroid percursors to estrone and estradiol. These choricarcinoma cells will be used with new hybrid strains to be developed to map genes for hormone synthesis to specific human chromosomes. The major techniques to be used to study hormone synthesis by the cells include: 1) Cell hybridization between hormone producing cells and non-endocrine mouse and human cells performed by fusing the cells and utilizing special selected media and cloning procedures to develop hybrid strains. Mutagenic agents will be used to alter hormone production in some parent and hybrid cells. 2) The modified strains will be examined for hormone production using chromatographic and radioimmunoassays for measurement of hormones and protein hormone subunits. Steroid synthesis will be examined by specific radioimmunoassays for specific steroids and by conversion of specific labeled hormone precursors into steroids. Subsequent studies will be performed utilizing complementation techniques utilizing different hybrid clones for correction of enzyme deficiencies in the steroid synthetic pathways. 3) Nucleic acid hybridization studies will be performed with hybrid strains utilizing labeled DNA complementary to the mRNA's for the alpha and beta subunits of hCG to identify hybrids containing structural genes for chorionic gonadotropin subunits. 4) Chromosomal analysis will be performed on hybrid cells utilizing enzyme assays and special staining techniques. These studies will be used to assign genes for specific hormones and subunits and enzymes in the steroid synthetic pathways to specific chromosomes. The control of hormone synthesis will be examined in the modified cell strains.